SOMATIC-CELL GENETIC AND BIOCHEMICAL-CHARACTERIZATION OF CELL-LINES RESULTING FROM HUMAN GENOMIC DNA TRANSFECTIONS OF CHINESE-HAMSTER OVARYCELL MUTANTS DEFECTIVE IN STEROL-DEPENDENT ACTIVATION OF STEROL SYNTHESIS AND LDL RECEPTOR EXPRESSION

We have isolated several non-leaky mutant Chinese hamster ovary (CHO) cell clones (M4, M19, and M21) requiring cholesterol and unsaturated f atty acid for growth. These mutants belong to the same complementation group as the mutant M1 cells previously reported from this laboratory . M19 cells revel fed to lipid prototrophy at very low frequency and w ere chosen as recipients to perform DNA-mediated gene-transfer experim ents using total human genomic DNAs. Biochemical characterization of t hese transfectant clones indicated that, unlike their parental M19 cel ls, they were able to exhibit activation of cholesterol biosynthesis a nd LDL receptor expression in response to sterol removal from the grow th medium. RNA blotting analysis indicated that these transfectants we re able to increase HMG-CoA synthase gene transcripts in response to s terol removal. From the genomic DNAs of a representative secondary tra nsfectant cell, we cloned a unique human DNA fragment (designated as h lambda 2) and showed that h lambda 2 is closely linked with the presu mptive human M1 gene.